Mårten Risling

Identification of a Neural Stem Cell in the Adult Mammalian Central Nervous System

New neurons are continuously added in specific regions of the adult mammalian central nervous system. These neurons are derived from multipotent stem cells whose identity has been enigmatic. In this work, we present evidence that ependymal cells are neural stem cells. Ependymal cells give rise to a rapidly proliferating cell type that generates neurons that migrate to the olfactory bulb. In response to spinal cord injury, ependymal cell proliferation increases dramatically to generate migratory cells that differentiate to astrocytes and participate in scar formation. These data demonstrate that ependymal cells are neural stem cells and identify a novel process in the response to central nervous system injury.

Mechanisms of blast induced brain injuries, experimental studies in rats

Traumatic brain injuries (TBI) potentially induced by blast waves from detonations result in significant diagnostic problems. It may be assumed that several mechanisms contribute to the injury. This study is an attempt to characterize the presumed components of the blast induced TBI. Our experimental models include a blast tube in which an anesthetized rat can be exposed to controlled detonations of explosives that result in a pressure wave with a magnitude between 130 and 260 kPa. In this model, the animal is fixed with a metal net to avoid head acceleration forces. The second model is a controlled penetration of a 2mm thick needle. In the third model the animal is subjected to a high-speed sagittal rotation angular acceleration. Immunohistochemical labeling for amyloid precursor protein revealed signs of diffuse axonal injury (DAI) in the penetration and rotation models. Signs of punctuate inflammation were observed after focal and rotation injury. Exposure in the blast tube did not induce DAI or detectable cell death, but functional changes. Affymetrix Gene arrays showed changes in the expression in a large number of gene families including cell death, inflammation and neurotransmitters in the hippocampus after both acceleration and penetration injuries. Exposure to the primary blast wave induced limited shifts in gene expression in the hippocampus. The most interesting findings were a downregulation of genes involved in neurogenesis and synaptic transmission. These experiments indicate that rotational acceleration may be a critical factor for DAI and other acute changes after blast TBI. The further exploration of the mechanisms of blast TBI will have to include a search for long-term effects.

Induction of VEGF and VEGF receptors in the spinal cord after mechanical spinal injury and prostaglandin administration.

Vascular endothelial growth factor (VEGF) is an angiogenetic factor that promotes endothelial cell proliferation during development and after injury to various types of tissue, including the central nervous system (CNS). Using immunohistochemical and in situ hybridization methods we have here demonstrated that VEGF and its receptors Flk‐1, Flt‐1 and Neuropilin‐1 mRNAs and proteins are induced after incisions in the rat spinal cord. The inducible enzyme for prostaglandin synthesis cyclooxygenase‐2 (COX‐2) is known to be upregulated after spinal injury, cerebral ischemia and to stimulate angiogenesis. To test the hypothesis that prostaglandins may be involved in the VEGF response after lesion we investigated whether intraspinal microinjections of prostaglandin F2α (PGF2α) alters VEGF expression in the spinal cord. Such treatment was followed by a strong upregulation of VEGF mRNA and protein in the injection area. Finally, by use of an in vitro model with cell cultures of meningeal fibroblast and astrocyte origin, resembling the lesion area cellular content after spinal cord injury but devoid of inflammatory cells, we showed that VEGF is expressed in this in vitro model cell system after treatment with PGF2α and prostaglandin E2 (PGE2). These data suggest that cells within a lesion area in the spinal cord are capable of expressing VEGF and its receptors in response to mechanical injury and that prostaglandins may induce VEGF expression in such cells, even in the absence of inflammatory cells.

Differential regulation of trophic factor receptor mRNAs in spinal motoneurons after sciatic nerve transection and ventral root avulsion in the rat.

After sciatic nerve lesion in the adult rat, motoneurons survive and regenerate, whereas the same lesion in the neonatal animal or an avulsion of ventral roots from the spinal cord in adults induces extensive cell death among lesioned motoneurons with limited or no axon regeneration. A number of substances with neurotrophic effects have been shown to increase survival of motoneurons in vivo and in vitro. Here we have used semiquantitative in situ hybridization histochemistry to detect the regulation in motoneurons of mRNAs for receptors to ciliary neurotrophic factor (CNTF), leukemia inhibitory factor (LIF), glial cell line‐derived neurotrophic factor (GDNF), brain‐derived neurotrophic factor (BDNF), and neurotrophin‐3 (NT‐3) 1–42 days after the described three types of axon injury. After all types of injury, the mRNAs for GDNF receptors (GFRα‐1 and c‐RET) and the LIF receptor LIFR were distinctly (up to 300%) up‐regulated in motoneurons. The CNTF receptor CNTFRα mRNA displayed only small changes, whereas the mRNA for membrane glycoprotein 130 (gp130), which is a critical receptor component for LIF and CNTF transduction, was profoundly down‐regulated in motoneurons after ventral root avulsion. The BDNF full‐length receptor trkB mRNA was up‐regulated acutely after adult sciatic nerve lesion, whereas after ventral root avulsion trkB was down‐regulated. The NT‐3 receptor trkC mRNA was strongly down‐regulated after ventral root avulsion. The results demonstrate that removal of peripheral nerve tissue from proximally lesioned motor axons induces profound down‐regulations of mRNAs for critical components of receptors for CNTF, LIF, and NT‐3 in affected motoneurons, but GDNF receptor mRNAs are up‐regulated in the same situation. These results should be considered in relation to the extensive cell death among motoneurons after ventral root avulsion and should also be important for the design of therapeutical approaches in cases of motoneuron death. J. Comp. Neurol. 426:587–601, 2000.

Effect of sciatic neurectomy on neuronal number and size distribution in the L7 ganglion of kittens

Abstract The number and size distribution of neurons in the L7 dorsal root ganglia were studied bilaterally 80 and 200 days after left-side sciatic nerve transection in kittens aged 1 week. The results show that about one-third to half of the neurons were lost on the operated side. Size spectra of surviving neurons showed a marked shift toward smaller sizes, most probably due to a combination of cell loss and atrophy or growth retardation of surviving neurons. The proportion of cells with intermediate sizes seemed to be increased, but the proportion of small cells was essentially unaltered. Both on the experimental and control sides the number of dorsal root axons exceeded the number of neurons. The absolute neuronal and dorsal root axon loss showed good agreement, but the proportion of lost neurons was larger than the relative decrease in dorsal root axon number. These results indicate that peripheral nerve transection results in a substantial loss of neurons in immature spinal ganglia, with no preference for a particular size class.

Expression of insulin-like growth factors and corresponding binding proteins (IGFBP 1-6) in rat spinal cord and peripheral nerve after axonal injuries.

Insulin‐like growth factors (IGFs) exert trophic effects on several different cell types in the nervous system, including spinal motoneurons. After peripheral nerve injury, the increased expression of IGFs in the damaged nerve has been suggested to facilitate axonal regeneration. Here we have examined the expression pattern of mRNAs encoding IGF‐1 and and ‐2, IGF binding proteins (IGFBPs) 1–6 in the rat spinal cord and peripheral nerve in three lesion models affecting lumbar motoneurons, i.e., sciatic nerve transection, ventral root avulsion, and a cut lesion in the ventral funiculus of the spinal cord. The expression was also studied in enriched Schwann cell and astrocyte cultures.

Impeded Interaction between Schwann Cells and Axons in the Absence of Laminin α4

The Schwann cell basal lamina (BL) is required for normal myelination. Loss or mutations of BL constituents, such as laminin-2 (α2β1γ1), lead to severe neuropathic diseases affecting peripheral nerves. The function of the second known laminin present in Schwann cell BL, laminin-8 (α4β1γ1), is so far unknown. Here we show that absence of the laminin α4 chain, which distinguishes laminin-8 from laminin-2, leads to a disturbance in radial sorting, impaired myelination, and signs of ataxia and proprioceptive disturbances, whereas the axonal regenerative capacity is not influenced. In vitro studies show poor axon growth of spinal motoneurons on laminin-8, whereas it is extensive on laminin-2. Schwann cells, however, extend longer processes on laminin-8 than on laminin-2, and, in contrast to the interaction with laminin-2, solely use the integrin receptor α6β1 in their interaction with laminin-8. Thus, laminin-2 and laminin-8 have different critical functions in peripheral nerves, mediated by different integrin receptors.

Localization of neurotrophin receptors in olfactory epithelium and bulb

We used in situ hybridization to localize trk, trkB and trkC mRNA, in rat and cat olfactory bulb. Expression of mRNA encoding truncated trkB receptors was seen in all layers, while only very modest full-length trkB expression could be detected. trkC hybridization was seen in all layers, most dense in the mitral cell layer. The localization of full-length tyrosine kinase trkB receptor in olfactory bulb and epithelium was examined with immunohistochemistry. trkB-like immunoreactivity was seen in the fila olfactoria, epithelium and in vitro, in olfactory sensory neurones. Since BDNF is expressed by olfactory sensory neurone target cells in the olfactory bulb, these data suggest that BDNF may act as a target derived neurotrophic factor in the primary olfactory system.

Experimental animal models for studies on the mechanisms of blast-induced neurotrauma

A blast injury is a complex type of physical trauma resulting from the detonation of explosive compounds and has become an important issue due to the use of improvised explosive devices (IED) in current military conflicts. Blast-induced neurotrauma (BINT) is a major concern in contemporary military medicine and includes a variety of injuries that range from mild to lethal. Extreme forces and their complex propagation characterize BINT. Modern body protection and the development of armored military vehicles can be assumed to have changed the outcome of BINT. Primary blast injuries are caused by overpressure waves whereas secondary, tertiary, and quaternary blast injuries can have more varied origins such as the impact of fragments, abnormal movements, or heat. The characteristics of the blast wave can be assumed to be significantly different in open field detonations compared to explosions in a confined space, such an armored vehicle. Important parameters include peak pressure, duration, and shape of the pulse. Reflections from walls and armor can make the prediction of effects in individual cases very complex. Epidemiological data do not contain information of the comparative importance of the different blast mechanisms. It is therefore important to generate data in carefully designed animal models. Such models can be selective reproductions of a primary blast, penetrating injuries from fragments, acceleration movements, or combinations of such mechanisms. It is of crucial importance that the physical parameters of the employed models are well characterized so that the experiments can be reproduced in different laboratory settings. Ideally, pressure recordings should be calibrated by using the same equipment in several laboratories. With carefully designed models and thoroughly evaluated animal data it should be possible to achieve a translation of data between animal and clinical data. Imaging and computer simulation represent a possible link between experiments and studies of human cases. However, in order for mathematical simulations to be completely useful, the predictions will most likely have to be validated by detailed data from animal experiments. Some aspects of BINT can conceivably be studied in vitro. However, factors such as systemic response, brain edema, inflammation, vasospasm, or changes in synaptic transmission and behavior must be evaluated in experimental animals. Against this background, it is necessary that such animal experiments are carefully developed imitations of actual components in the blast injury. This paper describes and discusses examples of different designs of experimental models relevant to BINT.

Increased trkb mrna expression by axotomized motoneurones

It has been previously demonstrated that members of the neurotrophin family may prevent death of motoneurones following axotomy. Here we report that the expression of mRNA for full-length trkB, which is a signal transducing receptor for BDNF and NT-4, increases transiently in spinal motoneurones after axotomy in adult rats. Thus, transection of the sciatic nerve resulted in markedly increased levels of trkB mRNA in axotomized motoneurones three days postinjury, while the levels had returned to normal within three weeks. The data support that neurotrophins may be important for survival and/or regeneration of motoneurones after axon lesion.